Leishmania are obligate intracellular parasites that are transmitted by a sand fly vector and cause a wide range of diseases, such as cutaneous (CL), mucocutaneous (MCL) and visceral leishmaniasis (VL). Over 12 million people currently suffer from leishmaniasis, and approximately 2 million are infected annually, making it a major global health problem and a neglected tropical disease. Recently, this infection is increasingly seen in dogs in the US as well as army personnel serving in Leishmania-endemic countries, Iraq and Afghanistan. Antimonials (GlucantimeTM and PentostamTM), amphotericin B, and miltefosine are used to treat leishmaniasis. However, the emergence of drug-resistant strains is rapidly increasing worldwide. Furthermore, these drugs are toxic and have poor patient compliance because many of them require systemic administration for periods ranging from 3-5 wks. Therefore, there is a strong need for new drugs that are safe, cheap, easy to administer and have broad-spectrum activity against different Leishmania. In the Yucatan Peninsula of Mexico, Mayan traditional healers have used Pentalinon andrieuxii root for the topical treatment of CL for many years indicating that P. andrieuxii contains antileishmanial molecules that could represent potential new drugs for leishmaniasis. We have found that a hexane extract of P. andrieuxii root (PARE) has potent antileishmanial activity both in vitro and in vivo. PARE kills Leishmania in vitro as efficiently as GlucantimeTM, and is effective against intracellular parasites. Our preliminary data show that topical treatment with PARE is also effective in limiting L. mexicana infection in mice. Work from our ongoing R21 project (Isolation of Novel Antileishmanial Molecules from P. andrieuxii root;AI076309) has already led to identification of one novel and several known sterols with leishmanicidal activity in PARE. More bioactive molecules are expected to be isolated from the plant root before the completion of this project in August 2010. The current proposal in response to RFA AI-09-026 is to expand our ongoing studies and to undertake preclinical testing on antileishmanial molecules isolated from the plant P. andrieuxii. In Aim 1, we will isolate and synthesize antileishmanial molecules from the different parts of the plant and identify their source(s). Aim 2 will use animal models to evaluate the safety and efficacy of these molecules in treatment of different forms of leishmaniasis caused by parasites that are sensitive or resistant to conventional drugs. Aim 3 will determine the mechanism(s) of action of these molecules. Our team is uniquely poised to perform the studies due to the complementary expertise in Leishmania pathogenesis (Satoskar), phytochemistry (Kinghorn), synthetic chemistry (Fuchs), mycology/endophytic fungi (Pearce) and antiparasitic drugs/pharmacology (Croft). Our studies should determine how bioactive molecules in P. andrieuxii mediate antileishmanial activity and provide information on their safety and efficacy to treat infections caused by Leishmania strains that are resistant to conventional treatment. Together, these data will lay the foundation for advancing future clinical studies on these molecules in the treatment of various forms of leishmaniasis.